Vegetative insecticidal proteins （VIPs）, produced during the vegetative stage of their growth in Bacillus thuringiensis, are a group of insecticidal proteins and represent the second generation of insecticidal trans-genes that will complement the novel δendotoxins in future. Fewer structural and functional relationships of Vip proteins are known in comparison with those of δ-endotoxins. In this study, both the maximum-likelihood methods and the maximum parsimony based sliding window analysis were used to evaluate the molecular evolution of Vip proteins. As a result, strong evidence was found that Vip proteins are subject to the high rates of positive selection, and 16 sites are identified to be under positive selection using the Bayes Empirical Bayesian method. Interestingly, all these positively selected sites are located from site-705 to site-809 in the C-terminus of the Vip proteins. Most of these sites are exposed and clustered in the loop regions when mapped onto its computational predicted secondary tertiary and a part of the tertiary structure. It has been postulated that the high divergence in the C-terminal of Vip proteins may not result from the lack of functional constraints, but rather from the rapid mutation to adapt their targeted insects, driven by positive selection. The potential positive selection pressures may be an attempt to adapt for the ＂arm race＂ between Vip proteins and the targeted insects, or to enlarge their target＇s host range. Sites identified to be under positive selection may be related to the insect host range, which may shed a light on the investigation of the Vip proteins＇ structural and functional relationships.
The cry gene family, produced during the late exponential phase of growth in Bacillus thuringiensis, is a large, still-growing family of homologous genes, in which each gene encodes a protein with strong specific activity against only one or a few insect species. Extensive studies are mostly focusing on the structural and functional relationships of Cry proteins, and have revealed several residues or domains that are important for the target recognition and receptor attachment. In this study, we have employed a maximum likelihood method to detect evidence of adaptive evolution in Cry proteins, and have identified 24 positively selected residues, which are all located in Domain Ⅱ or Ⅲ. Combined with known data from mutagenesis studies, the majority of these residues, at the molecular level, contribute much to the insect specificity determination. We postulate that the potential pressures driving the diversification of Cry proteins may be in an attempt to adapt for the ＂arm race＂ between δ-endotoxins and the targeted insects, or to enlarge their target spectra, hence result in the functional divergence. The sites identified to be under positive selection would provide targets for further structural and functional analyses on Cry proteins.
Jin-Yu WuFang-Qing ZhaoJie BaiGang DengSong QinQi-Yu Bao
